CN111218457A - Rice MIT2 gene and encoding protein and application thereof - Google Patents

Abstract

The present invention provides a riceMIT2The coding sequence of the gene and the coded protein thereof is shown as SEQ ID No.2, and the coding protein sequence thereof is shown as SEQ ID No. 3. The gene is expressed in a plurality of tissues. Experiments prove that MIT2 has the function of inhibiting rice tillering, so that rice plants become high. The invention also providesMIT2Mutant gene of gene and allelic mutant thereofmit2‑1They can make rice show that its plant height is changed into short, tillering is moderately increased, branch number is reduced, fertility is reduced, grain is reduced and glume is opened. The rice provided by the inventionMIT2Gene and mutant thereof in rice germplasmThe genetic improvement and breeding of resources has great effect, and is expected to regulate and control the formation of rice plant types and the tillering number, so that the plant types or the tillering numbers are directionally designed to improve the rice productivity.

Description

Rice and its production processMIT2Gene and its coding protein and application

Technical Field

The invention belongs to the field of genetic engineering, and particularly relates to riceMIT2Genes, proteins coded by the genes and application of the genes in inhibiting rice branches.

Background

Rice (1)Oryza sativa)Belongs to the gramineae rice genus, and is one of the most important grain crops. The tillering of rice is a very important agronomic trait, and one effective tillering corresponds to one ear of grain. Tillering is a key factor in determining rice yield. Rice tillering is not only an important factor related to rice yield, but also a core topic related to plant type in biological discussion. Several genes and Quantitative Traits Location (QTL) associated with tillering have been located. Therefore, the rice tillering regulation gene is further excavated, the tillering mechanism is clarified, and the rice tillering regulation gene has great significance for shaping rice plant types and improving rice yield.

The formation process of rice tillering is divided into two steps, i.e. the formation of tillering buds and the elongation of tillering buds. The axillary buds of each leaf of the rice can form a tillering bud (also called axillary bud and lateral bud), usually, only the axillary bud positioned on the non-extending internode at the base part of the stem can extend to form tillering, and the axillary bud positioned on the extending internode at the upper part of the stem can not extend generally but is in a dormant state. Therefore, the number of tillers of rice depends not only on the number of tillering buds formed, but also on the number of tillering buds capable of elongation. The formation of rice tillering is a complex biological phenomenon and is regulated and controlled by multiple factors such as genetic factors, plant hormones, external environment and the like. The hormones involved in the regulation of plant branches mainly comprise auxin, cytokinin, strigolactone and derivatives thereof, and the hormones have synergistic effect and jointly regulate the branch development of plants. The external environmental conditions including light, temperature, moisture, cultivation density, transplanting depth and plant nutrition also affect the differentiation and growth of rice tillering.

The first discovered key gene for regulating rice tillering isMOC1,MOC1Belongs to the GRAS family and encodes a transcription regulatory factor. Expressed in tillering buds, in axillary meristemsThe different stages of tissue vegetative growth and reproductive growth regulate the initiation and growth of the tillering bud. Rice mutantmoc1Almost completely loses tillering capability, only 1 main stem without tillering appears, and spikelets on the panicle are also obviously reduced, which indicates that MOC1 plays a positive regulation role in rice tillering and spike branching. Rice (Oryza sativa L.) with improved resistance to stressTB1The gene is a BRANCHED regulatory gene, TEOSINTE BRANCHED 1 (of maize)TB1) The similar sequences of (a) were cloned.

Research shows thatOsTB1Downstream of the Strigolactones (Strigolactones) signaling pathway, both genes encode putative transcription factors that have a basic helix-loop-helix DNA binding motif, designated the TCP domain.OsTB1Demonstration of genetic loci with maizeTB1Is a homologous gene. Overexpression of OsTB1 was found to reduce tillering numbersfc1The tillering of the mutant is increased, and the function of OsTB1 is lost. These results show that OsTB1 negatively regulates branching in rice, and in maizeTB1The genes function similarly. Multi-tillering mutant of riceteSeparating a negative regulatory factor of rice tillering by map-based cloning and transgenic verification as a materialTEHas a function of conservative cell cycle regulation. Experiments showed that there was a direct interaction between TE and MOC 1; the genetic experiment also proves thatTEAndMOC1on the same signal path. TE inhibits the tillering of rice by mediating the degradation of MOC 1. IPA1 protein can be used together with the rice tillering negative regulatorOsTB1The GTAC sequence of the promoter region of the gene is combined, and the combination can occur in the stem tip and the young ear.ostb1Andipa1the phenotype of the double mutant of (a) shows,ostb1can inhibitipa1Tillering phenotype ofipa1The tillering phenotype of the mutant may be due toIPA1Gene passageOsTB1Direct regulation, indicatingOsTB1May take part inIPA1Mediated transcriptional regulation of rice tillering.

Auxin (Auxin) is the first discovered plant hormone, is synthesized in the stem tip and young leaves of plants, and plays an important role in the growth and development process of the plants. The auxin inhibits the growth of tillering buds through apical dominance; auxin is mainly from top to bottomAnd (4) moving, transporting and directly conveying the tillering buds into the tillering buds to inhibit the growth and development of the tillering buds.YUCIs a key gene in the synthesis path of auxin and is mainly expressed in rice meristems and vascular tissues. The research finds that the content of the active ingredients in the active ingredients is high,YUCthe loss of (a) can cause the synthesis of auxin to be greatly reduced, so that the top advantages of the plant are lost, the rice tillering bud breaks dormancy, normal growth and development are started, and finally new tillering is developed.OsPIN1Is a gene encoding auxin transporter, is positioned in xylem parenchyma cells and regulates the transport of auxin into roots.OsPIN1The loss of function of (a) can cause the reduction of auxin transport capacity, so that the inhibition effect of the auxin on lateral bud growth and development is weakened, and the elongation of rice tillering buds is promoted.

Strigolactones are a class of sesquiterpene compounds identified as a novel plant hormone. The important breakthrough in the aspect of illustrating rice tillering is to find strigolactone which is mainly synthesized in roots, is generated by carotenoid metabolism, is transported from bottom to top and inhibits the growth of tillering buds. Strigolactone is a novel plant hormone which can inhibit the development of lateral branches besides auxin and cytokinin. At present, strigolactones have been isolated from many plants, and genes involved in strigolactone synthesis and signal transduction have also been sequentially found in higher plants such as Arabidopsis thaliana, pea, petunia, rice, and lower plant mosses. In rice, several genes have been shown to be involved in the strigolactone-mediated collateral development process.D17/ HTD1AndD10the encoded carotene-cleaving dioxygenases 7 (CCD 7) and 8 (CCD 8) are two key enzymes of the strigolactone synthesis pathway. Researchers cloned from Arabidopsis thaliana mutants with increased axillary branchesMAX1ToMAX4The gene(s) is (are),MAX1，MAX3andMAX4both participate in the biosynthesis of strigolactone, and MAX2 participates in the signal transmission of strigolactone. In riceD3Encodes a member of an F-BOX protein family, is a homologous protein of Arabidopsis MAX2, participates in protein ubiquitination and mediates protein degradation by forming an SCF (Skp 1-Cull-in-F-BOX) protein complex, and is a key gene of a strigolactone signal transduction pathway. Rice geneD27AndD14also demonstrated to be members of the strigolactone synthesis and signal transduction pathways.D27Encodes a novel iron-containing protein and influences the polar transport (PAT) of auxin, and participates in the synthesis of strigolactone.D14/ D88/ HTD2Encodes a hydrolase/esterase, which is a signaling receptor for strigolactones. The rice mutants are shown as a multi-tillering and dwarf phenotype, the monolaurate controls the development of axillary buds, and the monolaurate, auxin and cytokinin jointly determine the plant type of rice.

A large number of physiological and biochemical data indicate that there is a certain relationship between hormones. Auxin inhibits the formation of rice tillers by promoting the synthesis of strigolactones. The strigolactone signal molecule also inhibits the auxin signal pathway and another unknown feedback regulation signal through a feedback pathway, and both signal pathways can regulate the biosynthesis of the strigolactone in plastids. The strigolactone and the auxin can mutually regulate the content of each other in the plant body according to environmental signals, and form a dynamic feedback cycle to jointly regulate the branches of the plant. Although there have been a lot of studies on the regulation of tillering of rice by plant hormones, their specific interrelations and how they cooperate to regulate the growth and development of tillering buds require further studies.

Disclosure of Invention

The invention aims to provide riceMIT2Gene and its coded protein and application.

The invention takes rice Nipponbare as a material to carry out EMS mutagenesis, and hopes to screen key genes which can regulate and control rice tillering and influence rice yield. The inventors have made an effort to screen rice mutants having a moderately increased number of tillers. Because excessive tillering is unfavorable for high yield of rice, most nutrition is used for vegetative growth of rice, and reproductive growth is not favorable. From the mutant library, the inventors screened out a plurality of mutants with moderate tiller number, named as Moderately Incorporated Tillers (MIT), and obtained mutants were named as MIT1, MIT2 … …, and the like, respectively, in the order of discovery. The mutant genes of the mutants are different, and the functions of the mutants in the rice tillering regulation process are different. The present invention has been studied with respect to the MIT2 gene and its function. The inventor finds that EMS-mutagenized rice mutants have the mutant MIT2 with half short and moderately increased plant tillering number, and the map-based cloning and function complementation experiments find that the mutation of the MIT2 gene causes the plant to become short and the moderately increased tillering number. The MIT2 gene encodes a protein of unknown function. The invention will explain the function of MIT2 in the rice tillering process.

The present invention firstly provides a rice MIT2 protein having:

1) an amino acid sequence shown as SEQ ID No. 3; or

2) Protein which is derived from the protein 1) and has equivalent activity and is obtained by substituting, deleting and/or adding one or more amino acids in the amino acid sequence shown in SEQ ID No. 3.

The present invention provides a gene encoding rice MIT2 protein, having:

1) a nucleotide sequence shown as SEQ ID No. 2; or

2) The nucleotide sequence shown in SEQ ID No.2 is substituted, deleted and/or added with one or more nucleotides; or

3) Nucleotide sequences which hybridize under stringent conditions with the DNA sequences defined in 1).

The invention provides a biological material containing a gene for encoding rice MIT2 protein, wherein the biological material is a plasmid, a vector, a host bacterium or a transformed plant cell.

The invention provides the application of any one or more of the following rice MIT2 protein, the encoding gene thereof or the biological material containing the encoding gene thereof:

(1) the application in preparing transgenic plants;

(2) the application in rice germplasm resource improvement;

(3) the application in maintaining rice fertility;

(4) the application of the rice grain enlargement agent in enlarging rice grains;

(5) application for increasing rice plant height

(6) The application in inhibiting rice tillering.

In a second aspect, the invention provides a rice MIT2 gene mutant gene, which is characterized in that the base T at the 2413 th position of a CDS on the 8 th exon of a rice MIT2 gene is deleted, and the sequence of the CDS is a nucleotide sequence shown in SEQ ID No. 2.

In a third aspect, the invention provides an allelic mutant of the rice MIT2 gene mutant gene, wherein the mutation site of the allelic mutant occurs at the splicing site of the 3 'end of the first intron, the original AG is changed into AA, and the first two bases AG at the 5' end of the second exon are cut off.

The invention provides a biological material, which contains the rice MIT2 gene mutant gene or the allelic mutant, and the biological material is a plasmid, a vector, a host bacterium or a transformed plant cell.

In a fourth aspect, the invention provides any one or more of the following applications of the rice MIT2 gene mutant gene or the allelic mutant or a biological material containing the same,

(1) the application in crop improvement breeding and seed production;

(2) the application in reducing the height of crop plants;

(3) the application in increasing the tillering number of rice;

(4) the application in reducing rice fertility;

(5) the application in cultivating rice with reduced grains;

(6) application in improving rice yield.

In the application, the plant height of the crops is reduced to 50-60% compared with the wild type. In the examples of the present invention, it was found that the phenotype of the mutant gene of MIT2 gene of rice or the allelic mutant is slightly short in the plant height in the mature period and 60% of the wild type.

In the application, the tillering number of the rice is increased by 2-3 times compared with that of a wild type. In the embodiment of the invention, the rice MIT2 gene mutant gene or the allelic mutant is found to increase the tillering number of rice by 2 times of that of the wild type.

The invention has the advantages that: the present invention provides riceMIT2Gene (nucleus)The nucleotide sequence is shown as SEQ ID No.1, the nucleotide sequence of the coding region is shown as SEQ ID No. 2) and the protein coded by the nucleotide sequence (the amino acid sequence is shown as SEQ ID No. 3). The MIT2 gene mutant of rice and the allelic mutant of the mutant gene have deletion or mutation of a plurality of genes compared with the wild type, which are obtained by EMS mutagenesis, the molecular detection can be carried out by adopting agarose electrophoresis commonly used in laboratories, namely, the identification can be realized, and special detection technology and method are not needed. Will be over-expressedMIT2Gene function complementary vector and self promoter drivenMIT2Genome complementary vector mediated transformation by agrobacteriummit2The phenotype of the complementary transgenic plant is completely recovered to normal by the callus of the mutant, and the tillering number and the plant height are consistent with those of the wild type, which indicates thatMIT2The gene has the function of inhibiting rice tillering, is expected to regulate and control the formation of rice plant types so as to directionally design the plant types, so that the rice productivity is improved, the screening effect in the improvement of rice germplasm resources is obvious, and the economic value is huge.

Drawings

FIG. 1 shows the phenotype of wild-type Nipponbare and mutants. (A) Performing in a mature period; (B) a tillering phenotype; (C) the mutant has high-node tillering; (D) ear type; (E) expressing the branches and stalks of the spike part; (F) a grain phenotype; (G) counting the plant height; (H) and counting the tillering number.

FIG. 2 isMIT2 Gene mapping and Structure. (A) Coarse positioning; (B) fine positioning; (C) map of MIT2 gene.

FIG. 3 is a drawing showingMIT2And (4) verifying the functional complementation of the genes. (A) Overexpression of MIT2 complemented the mutant phenotype; (B) MIT2 genome driven by self promoter can complement mutant phenotype; (C) counting the plant height; (D) and counting the tillering number.

FIG. 4 is a drawing showingMIT2Expression in various tissues of rice.

FIG. 5 shows the GUS staining results of rice tissues. (A) Internode; (B) a leaf sheath; (C) young ears; (D) small ears; (E) glumes; (F) a stem base; (G) tillering buds; (H) and (4) root.

FIG. 6 is the subcellular localization of MIT2 protein.

FIG. 7 is the structural diagram of vector pCAMBIA 1305.1AP FH-N.

FIG. 8 is a schematic diagram of pCAMBIA 1305.1-GFPC structure.

Detailed Description

The following examples are intended to illustrate the invention but are not intended to limit the scope of the invention. Modifications or substitutions to methods, procedures, or conditions of the invention may be made without departing from the spirit and scope of the invention.

Unless otherwise specified, the technical means used in the examples are conventional means well known to those skilled in the art; all reagents used in the examples are commercially available unless otherwise specified.

Example 1 acquisition and phenotypic analysis of mutants

Through EMS chemical mutagenesis of japonica rice variety Nipponbare, a tiller moderate-increase mutant is obtainedModerately Increased Tiller-2(mit2) And allelic variants thereofmit2-1。mit2Mutation is base deletion, and allelic mutant thereofmit2-1Has similar dwarfing and multi-tillering phenotype and is mutated into base replacement.

Phenotypic analysis showed that the mutant was altered in many ways compared to the wild type (FIG. 1). Rice (Oryza sativa L.) with improved resistance to stressmit2The height of the mutant plant is shorter than that of the wild type plant, the plant height in the mature period is slightly shorter, about 60 percent of that of the wild type plant, and the equal proportion among all the internodes is shortened to a certain extent;mit2the tillering degree of the mutant is increased by about twice of that of the wild type. Mutantsmit2And allelic mutants thereof have fewer branches, reduced fertility, smaller grains and open glumes.

Example 2 RiceMIT2Gene acquisition and functional complementation verification

1. Rice (Oryza sativa L.) with improved resistance to stressMIT2Obtaining of genes

According to the inventionMIT2The gene is obtained by mutant by adopting a map-based cloning methodmit2Obtained by cloning. Homozygous mutantmit2Crosses with the phenotypically normal and highly polymorphic indica variety Dular, all crosses F₁The middle individuals show normal plant height and tillering number. F₂Obtaining segregating population, carrying out genetic analysis and gene determinationA bit. Analysis of the strain with character segregation in F2 generation showed that the segregation ratio of wild type and mutant was in accordance with the 3:1 genetic segregation ratio, thus showing that the mutant character is controlled by a pair of recessive genes.

In order to control the position of the mutant gene with moderate tillering increase in a positioning way, F1 generation self-bred F is utilized to construct₂Segregating the population as the population to be mapped, and selecting l0 strain F by BSA method₂Mutant individuals construct a DNA pool, and the candidate gene is located on chromosome 9 by using 170 Indel markers uniformly distributed on 12 rice chromosomes (A in FIG. 2). In order to clarify the location of the candidate gene on chromosome 9, 50 individuals were selected and mapped using molecular markers (Table 1) developed in the laboratory and distributed uniformly throughout the chromosome, indicating that the candidate gene was mapped between the R9-1 and R9-2 markers. For fine localization of the gene of interest, F₂The mapping population is expanded to 6380 dwarf tillering moderate increase individuals, 6 pairs of new Indel markers are developed, and finally the target gene is mapped between the markers M3 and M4 (B in figure 2). The physical distance between these two markers is about 1.2 Mb, containing 13 candidate genes. Sequencing the DNA sequence to find out the geneLOC_Os09g06560A mutation occurs.

LOC_Os09g06560The total length of the genome DNA is 6325 bp (shown as a sequence in SEQ ID No. 1), the genome DNA comprises 12 exons and 11 introns, the total length of a coding region is 3378 bp (shown as a sequence in SEQ ID No. 2), and 1125 amino acids (shown as a sequence in SEQ ID No. 3) are coded. 13 pairs of primers are designed for sequencing, and the analysis of the gene sequencing result shows that,mit2the mutation site is located on the 8 th exon, and the 2413 rd base T of CDS is deleted, so that the translation process is shifted from the 805 th amino acid. Allelic variants thereofmit2-1The mutation site occurs at the splicing site of the 3 'end of the first intron, the original AG is changed into AA, sequencing finds that the mutation causes the splicing error of the gene, and the first two bases AG at the 5' end of the second exon are cut off (C of figure 2)）。

2、MIT2Functional complementation verification of genes

The invention is toMIT2Cloning of genes into plant expression vectorspCAMBIA 1305.1AP FH-NAndpCAMBIA 1305.1- GFPC(the vector structure is shown in FIGS. 7 and 8 in this laboratory), and overexpression was constructed for the purpose of performing the function complementation experimentMIT2Gene function complementary vector (pCAMBIA 1305.1-Actinpro-FLAG-HA-MIT2CDS) And driven by their own promotersMIT2Genome complementary vector (pCAMBIA 1305.1-MIT2pro-MIT2genomic DNA-GFP) The obtained transgenic plants were designated B267 and B270 with FLAG and GFP tags, respectively.

Over-expressionMIT2Gene function complementary vector (pCAMBIA 1305.1-Actinpro-FLAG-HA-MIT2CDS) The construction of (1): in thatMIT2The 5 'end of the full-length CDS is introduced into NcoI site, the 3' end is introduced into SpeI site, and the fragment length is 3378 bp. The primers used were 09g06560S1 SPF: CGAACGATAGCCATGGCCATGATATTTCAGCTAAGA AATGCG and 09g06560S1 SPR: GGTAGGATCCACTAGTCACTTTCACCT TCTTGCTGTTAGC are provided.

Driven by self-promotersMIT2Genome complementary vector (pCAMBIA 1305.1-MIT2pro-MIT2genomic DNA-CGFP) The construction of (1): 2460bp upstream of the ATG of the MIT2 gene is defaulted to be the promoter of the MIT2 gene, an EcoRI site is introduced at the 5 'end position of the promoter, and a KpnI site is introduced at the 3' end before the termination codon TAA of the MIT2 genome sequence. The MIT2 promoter sequence 2460 bp; the MIT2 genome sequence is 5300bp, and the fragment is 7760bp in total length. The primers used for the promoter were: promF: CCATGATTACGAATTCTGCTATGCCGTTAGGTA GCAC and promR: CCCTTGCTCACCATGGTACCCTGAAATATCATTGCTAACCATCA; the sequences of the primers used in the genome are GDNAF: CAATGATATTTCAGGGTACCATGATATTTCAGCTAAGAAATGCG and GDNAR: CCCTTGCTCACC ATGGTACCCACTTTCACCTTCTTGCTGTTAGC. Recombining both fragments togetherpCAMBIA1305.1-CGFPAnd (4) removing.

The two constructed expression vectors are transferred into agrobacterium EHA105 by an electric shock method, and the ricemit2The seed induction callus of the mutant knot is used as a receptor material, and the agrobacterium-mediated transformation method is used for rice transformation. The obtained transgenic plant has complete phenotypeThe mutant is completely restored to be normal, the tillering number and the plant height are consistent with those of a wild type (figure 3), the number of the mutant branches, the fertility and the hard shell opening condition are also restored to be the phenotype of the wild type, and the fact that the mutant phenotype can be complemented by over-expression of MIT2 and the mutant phenotype can be complemented by an MIT2 genome driven by a self promoter is shown. The above results prove thatLOC_Os09g06560Is the target geneMIT2。

Example 3 RiceMIT2Gene expression patterns

To clarifyMIT2Extracting RNA of different tissues (seeds, roots, stems, leaves, leaf sheaths, scions, flowers, tillering buds, seedlings and radicles) of japonica rice varieties Nippon japonica, carrying out reverse transcription to obtain cDNA, detecting the expression level of the gene in each tissue of rice by using a Real-time PCR method by using a rice Ubiquitin gene as an internal reference, and displaying the resultMIT2The gene was expressed in all the above tissues of rice, with the highest expression level in young roots, higher expression level in young ear seeds and seedlings, and relatively lower expression level in leaf sheaths and tillering buds, as well as in seeds and mature roots (FIG. 4).

Example 4 GUS staining results in Rice tissue

MIT2promoter::GUSVector construction:MIT2the promoter region is obtained by PCR amplification, BamHI site is introduced at 5 'end, NcoI site is introduced at 3' end, the fragment length is 2460bp, and recombination is carried outpCAMBIA1305.1In the BamHI and NcoI sites. The forward primers used were: CGGTACCCGGGGATCCTGCTATGCCGTTAGGTAGCAC, respectively; reverse primer: CTCAGATCTACCATGGGAAATATCATTGCTAACCATCA are provided. A transgenic plant is obtained by a method of infecting rice calluses with agrobacterium tumefaciens and is used for analyzing a gene tissue expression mode.

Detection using GUS (β -glucuronidase) gene as reporter geneMIT2Spatial distribution of gene expression. GUS activity was detected in internodes, leaf sheath, stem base and tillering bud. GUS was hardly active in mature roots, anthers (FIG. 5).

Example 5 subcellular localization of MIT2 protein

Self promoter driven constructed in example 2MIT2The genome complementary vector is fused with GFP label, and the obtained transgenic plant B270. During the germination period of the transgenic seeds, the subcellular localization of GFP at the root tip of the rice is observed by a laser confocal microscope, and the result shows that the MIT2-GFP protein is mainly expressed in a nucleus at the root tip part (figure 6).

The foregoing is only a preferred embodiment of the present invention, and it should be noted that, for those skilled in the art, various modifications and decorations can be made without departing from the technical principle of the present invention, and these modifications and decorations should also be regarded as the protection scope of the present invention.

Sequence listing

<110> institute of crop science of Chinese academy of agricultural sciences

<120> rice MIT2 gene and its coding protein and application

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catcgccaac gcccgctcca gcctttcggc aacggagcct cctccacgcc acgtcagcct 60

cttcggtgac gacctccttg cctccaccgc tgccaaacgc gccggcctcg tcgtccgacc 120

gtcacttgca ttgtcctcgt cctccgccat cgaccatgac cgtgcttcgt catgtcgccc 180

tgtgccgccc cgctcctcca ccgggccgcc gtcaccgcga gcgccattgc tgttgaggga 240

ttcggcaccg ccggagttgc ccgctcccaa tcttgctgca ggagggctag atgcgcatcc 300

ggcgtggcca aattggccgc gccgagtact ctacccaccg gtgcatgcca ctccacctcc 360

cgtgccgaga agaagcgtca ttaacttgtt ttgcctgcca ccgccttctt ggttggccgc 420

tcggtttccg acggtggcga tgcagcaaga aagagtgagg aaggtggcgg tgttttgccg 480

ccagccgccc gcgcaagagc aatgctggga caaggaaaaa aaaatccaaa agtatataac 540

gtgctggact ttgactaaaa agcacaattt tttttattgt gatggttagc aatgatattt 600

cagctaagaa atgcgatagt taaacatata ttccttttga gtttattttt gaaaatatcc 660

ctaccttacc tccgccactg cggccgtgaa attcggtgta tttttcccgc ggtactccac 720

caaaagcttc ccgccaaacg tcctttcccg cgcgggggga cgcgccaaaa cccaccaaaa 780

tgtctgcaac caaaacccca ccccgcgtat cgtcaatctc tggaaacccc taacaaattc 840

ctgaacaccc gccgccacca tagcttcagc ctcgtgggtg aagaatctct cgtcgtcgtc 900

gtcggtgtca ccatgaaagg gcgcgcggtg aagctccgag aggcgcacaa ggccggctcg 960

ccagtcttct gctccgttgc gtggggccaa ggcgggcagc atgtcgtcac cgcatccgcc 1020

gccgacgtgg ccatcctcat ccatgacgcc gctgcggtcg ccgccgccgg tggccggagc 1080

tcgggctccg cggctgcggc ggcgctttcc acgatccggc ttcacaagga tggcgtcacg 1140

gcgctcgccg tcgcgccggg ctccggcgcg tcgctggcgt ccggctccat cgatcactcc 1200

gtcaagttct gttctttccc aggttcttag atctgactgc cccggatgca atttctccta 1260

atctccgtca tttccacggc tatatttgtg aaattttgct tcggatttcc ttccagaggg 1320

ggtgttccag agcaatatcg cccggttcac cctgccgatc cggtcactgg ccttcaacaa 1380

gaaggggact ctgctggcgg cggccggaga cgacgacggc atcaagttga ttgccaccat 1440

cgacaacacc atctccaaag tgctcaaggg ccacaaggga tcggtaaccg ggttgtcttt 1500

cgatcccaga aacgattatt tggcatcaat tgacaccttc ggcacagtca tcttctggga 1560

tctctgcacg gggactgaag cccgtagtct gaagcggatt gcgccgacat ttggttcaga 1620

ccactcaatc aacaatgccc tgtgctggag ccctgatggg cagttccttg ctgttccggg 1680

attgaggaat aatgtggtca tgtatgatag ggacaccggt gaggaggtgt tcactctgaa 1740

aggggagcat gagcaaccag tgtgtagtct ctgctggtct ccaaatggga ggtacctagt 1800

cactgctgga ttggataagc aggttctgat ctgggatgtg aagtcaaagc aggatgttga 1860

gaggcagaag ttcgatgaaa ggatatgtag cttggcttgg aaacctgaaa gtaatgctgt 1920

agcagtgatc gacgtaactg gcagatttgg catttgggaa tcggtcatcc cgtcgacttt 1980

gaaatcgccc acagagggtg cacctgacct gaactctact aaggttcctt tgtttgatga 2040

cgaggatgat gaggagaggc cgagtacctc tggtggactg gatgatgatg atgatgatga 2100

aagtcttggt gaattaggtc cattcaacca caagagattg aggaggaagt caacctatca 2160

tgatcactca aatggagata gtgaagatga ggatctgata cttcagatgg agtcacgcaa 2220

gagaatgaaa gatacacata gagataacaa ggaggttgct gataaggcaa taggtgattc 2280

agcaacttca gtaagactgg ttacagcaag aatgcaaact gcatttcagc ctgggtccac 2340

accacctcaa cctggcaagc gaaatttcct tgcctacaat atgcttggaa gtatcactac 2400

tatcgaaaat gaggggcatt cacatgtaga ggtaaaatct tctcaccctc tatcttataa 2460

gccattgtat cctctacttg tttgcagctt ggatgtgaat aaaccatccg aagttacttt 2520

gtttttcagg tagacttcca tgacaccgga agaggtccta gagttccttc gatgactgat 2580

tattttggtt tcacaatggc tgcactgaat gaatcaggaa gtgtctttgc aaatccatgc 2640

aagggtgaca agaatatgag cactcttatg taccgccctt tcagtagttg ggcaggcaac 2700

agtgaggtaagttaactaaa tgaaattgtt gtttgccagc ttctgagata gggttgaagt 2760

ttaccacttt ggtgcttact ggactaattt gaaaattact tagtggtcaa tgaggtttga 2820

gggagaagaa gtgaaggctg tggctgttgg tgttggatgg gtcgctgcag ttacaacttt 2880

aaattttctg cgcattttca cagagggagg gttgcaggtt ttgtaacttc ctcaagcttt 2940

gtatttaagt ccttttttcc tgatgaatat acttgttagt cagtttgtga agttcatttt 3000

tttcctgaaa gaaagctgac tccttattag ttccgaaata cccaaaaaac caggttgtgc 3060

ctggtaatca gtttgtgaag ttcatttttt tcctgaaaga aaactgcctc cttattagtt 3120

ccgtaatacc caaaaaacca ggttgcttta gtttttttca gtgctatcta ctagagcttt 3180

cataaggcca aaatagctgt gaagtattaa caaaagaatt tatagtattt taaaaacaat 3240

taatgttata gaagaaaatt agattcatgc gactctcgac atgtttccaa ttaaatttat 3300

tggtcaagga aacattctat actaccaaca tttaaatgct gaaaccattt tttatttcta 3360

catccaatgt acagccgaaa ttgtcccgcc ctaaccatca gaagcacaga aattgttgta 3420

ctatgtaacc actcatgaat tataacttgt ggggcacgaa ttataacttt gactttttgc 3480

actcaagaaa ataattcagg gaatgcactt tccatgtaga tactagtaac ttttatcacc 3540

atttctcacc aagaagtggt ctacggaaca catttacatt tttagatcat taaaaatgct 3600

gttcatgaca ttggtcgtga cttacatagc aaagccaaac tacatattac taatcttgga 3660

tggtttttct tgatacatgc gagctggggg ttacataagc attcatgaat ctgatttaca 3720

tcattccctg tttgttactt gaagtgggaa aaattcaatt taatttactc attttatatt 3780

gtacttttac ctagcatgtc ttgattttca gatgcatatc ctcggtccgt ggcccaatta 3840

aaaactcttt aagttttgtc aaacacatct cttgattttc agatgcatat cctctcagtc 3900

ggtggcccag tggttactgc ggcaggccat ggagatcagc tagcgattgt gtctcatgct 3960

tcagattgtc tttcatcagg agaccaggta ttcagtatat gttcgtttga cttttggagc 4020

tttgtgtagt gatgatatat gttgtagcta gctatcactt atcagattgt cagcacctcc 4080

tagtgtacca tgattccaat cgtgtgaacc aattcacagt acatatgaac tgatggttga 4140

aatggtacta caagattctg aattgaaaca tggatttgtt catgattaac cctttcaggt 4200

gctggatgtt aaagtactga agatatctga atgtgctcaa tcattgtcca gccgacttgt 4260

tttaactcct gcctctaaat tatcttggtt tggtttcagt gagaatggtg aacttagttc 4320

ctttgattcc aaggtatgac caatgaacac tgtcaacaat tcaacatatc tttagttata 4380

tcatgggagt tttaaactga atattgttat ttctagggaa tactgagggt cttttctggt 4440

caattcggtg gaagctggat tccaatattt aggtatagta tctgcctgac aggttatgca 4500

ttttactact accaaagcac acttaataat tatatccaac tgcagttcaa tcaaggcaag 4560

aaaatccgaa gatgaaagcc actgggtggt gggcttagat gctaataata tattctgcat 4620

tctatgcaag tccccggagt cctatccaca ggtatgccta aatgcaactt ctcatgtgta 4680

caacttttga gtgttgatgt tccaacaaaa atagtacttc tatatgcaat tctcaaatat 4740

tgaaatttgt ttgtcgtttg acgtctttga ctgatcttat caggtgatgc ccaaacctgt 4800

tttgacaata cttgagctgt catttcctct tgcatcatct gaccttggtg ccaatagttt 4860

ggaaactgaa ttcatgatga ggaaactgca tctctcacag gtgcactgtg tttctttcag 4920

ttttgaaggg cagtatctgt gatattgttt ttaatcgttg ctgtttatct cagattcaaa 4980

agaaaataga agaaatggct gctttgggtc tggacacgat tgcattagat gatgaagcat 5040

tcaacatgga ggctgcactt gaccggtgca tcttgaggct catctccagc tgctgcaatg 5100

gtacaagact tgaactatgc tttttaccca taaatagtgc tttgcattaa attatttctt 5160

gttttatttc ttgatttcaa aaggtgataa gcttgtacga gctaccgagc ttgcaaaatt 5220

actaacactg gagaagtcaa tgaagggagc attgatgctt gttacacgct taaaacttcc 5280

catattgcaa gaaaggttca gtgccatact cgaggtgaaa ttcacaaccc ttatactgaa 5340

cctgcgtact tttagcactc taaaaatggc acattcaata tttaagaatt tatcatatgt 5400

tcaactgaag gttatgcaat ggcttcttgt aggagatgat gctaaacaat gcaaaaattg 5460

ccaatacatc tggtgttttc tccaatagta atacaaacta ctcaccatca ccagcgttga 5520

gcactcaagc agtcccacca gctaaggttg tgcaaaatgg aaacagcttg aagttaccta 5580

cattgcctaa actgaatcct gccgcccaac gaagcaatcc aactgaatca aacaaggcag 5640

aggtagaaca agcagacaat ttgaaagaaa tcagtacaaa ggtttcacct gcacaaactc 5700

cgttagttaa aattccaaaa aacagtgaaa tgggtgtaaa aacgaagaaa gataatgatg 5760

gagcatcaca tgcaactaca gttgatcaga acccaaaggg aggcagtggt caggttggcc 5820

ttaaaaacaa gagcgtcgat agctgcaatg gtgtacagcc tcagcggcca gttaacccct 5880

tcgcgaaatc ctcatcaagc aaagaacagc catcatccct ttttgattcc atcaagaaga 5940

tgaaggtcga aaatgagaag gttgacaaag ctaacagcaa gaaggtgaaa gtgtaactgc 6000

ttgtcattat ctcatgatct tcgggagcaa accagcagtt aggtatcctc cacctttact 6060

gacatatctg tgcttatatt tttcatttca gttgtgataa tgtaggtgaa aatcactatt 6120

tacttcttga gctacgtttt tgtgtatggc ttcagagctt aatttctagt tatgaggtgg 6180

tacacaatgt atgatagtaa acctgtttag atagtttgct tgttcctttt gttgattgct 6240

ttgatataca attacatttt cgtctgatta tatttaggat ttatgtatct cctacagata 6300

tataatggag tgcgcaggtt ggctg 6325

<210>2

<211>3378

<212>DNA

<213> Artificial Sequence (Artificial Sequence)

<400>2

atgatatttc agctaagaaa tgcgatagtt aaacatatat tccttttgag tttatttttg 60

aaaatatccc taccttacct ccgccactgc ggccgtgaaa ttcggtgtat ttttcccgcg 120

gtactccacc aaaagcttcc cgccaaacgt cctttcccgc gcggggggac gcgccaaaac 180

ccaccaaaat gtctgcaacc aaaaccccac cccgcgtatc gtcaatctct ggaaacccct 240

aacaaattcc tgaacacccg ccgccaccat agcttcagcc tcgtgggtga agaatctctc 300

gtcgtcgtcg tcggtgtcac catgaaaggg cgcgcggtga agctccgaga ggcgcacaag 360

gccggctcgc cagtcttctg ctccgttgcg tggggccaag gcgggcagca tgtcgtcacc 420

gcatccgccg ccgacgtggc catcctcatc catgacgccg ctgcggtcgc cgccgccggt 480

ggccggagct cgggctccgc ggctgcggcg gcgctttcca cgatccggct tcacaaggat 540

ggcgtcacgg cgctcgccgt cgcgccgggc tccggcgcgt cgctggcgtc cggctccatc 600

gatcactccg tcaagttctg ttctttccca gagggggtgt tccagagcaa tatcgcccgg 660

ttcaccctgc cgatccggtc actggccttc aacaagaagg ggactctgct ggcggcggcc 720

ggagacgacg acggcatcaa gttgattgcc accatcgaca acaccatctc caaagtgctc 780

aagggccaca agggatcggt aaccgggttg tctttcgatc ccagaaacga ttatttggca 840

tcaattgaca ccttcggcac agtcatcttc tgggatctct gcacggggac tgaagcccgt 900

agtctgaagc ggattgcgcc gacatttggt tcagaccact caatcaacaa tgccctgtgc 960

tggagccctg atgggcagtt ccttgctgtt ccgggattga ggaataatgt ggtcatgtat 1020

gatagggaca ccggtgagga ggtgttcact ctgaaagggg agcatgagca accagtgtgt 1080

agtctctgct ggtctccaaa tgggaggtac ctagtcactg ctggattgga taagcaggtt 1140

ctgatctggg atgtgaagtc aaagcaggat gttgagaggc agaagttcga tgaaaggata 1200

tgtagcttgg cttggaaacc tgaaagtaat gctgtagcag tgatcgacgt aactggcaga 1260

tttggcattt gggaatcggt catcccgtcg actttgaaat cgcccacaga gggtgcacct 1320

gacctgaact ctactaaggt tcctttgttt gatgacgagg atgatgagga gaggccgagt 1380

acctctggtg gactggatga tgatgatgat gatgaaagtc ttggtgaatt aggtccattc 1440

aaccacaaga gattgaggag gaagtcaacc tatcatgatc actcaaatgg agatagtgaa 1500

gatgaggatc tgatacttca gatggagtca cgcaagagaa tgaaagatac acatagagat 1560

aacaaggagg ttgctgataa ggcaataggt gattcagcaa cttcagtaag actggttaca 1620

gcaagaatgc aaactgcatt tcagcctggg tccacaccacctcaacctgg caagcgaaat 1680

ttccttgcct acaatatgct tggaagtatc actactatcg aaaatgaggg gcattcacat 1740

gtagaggtag acttccatga caccggaaga ggtcctagag ttccttcgat gactgattat 1800

tttggtttca caatggctgc actgaatgaa tcaggaagtg tctttgcaaa tccatgcaag 1860

ggtgacaaga atatgagcac tcttatgtac cgccctttca gtagttgggc aggcaacagt 1920

gagtggtcaa tgaggtttga gggagaagaa gtgaaggctg tggctgttgg tgttggatgg 1980

gtcgctgcag ttacaacttt aaattttctg cgcattttca cagagggagg gttgcagatg 2040

catatcctct cagtcggtgg cccagtggtt actgcggcag gccatggaga tcagctagcg 2100

attgtgtctc atgcttcaga ttgtctttca tcaggagacc aggtgctgga tgttaaagta 2160

ctgaagatat ctgaatgtgc tcaatcattg tccagccgac ttgttttaac tcctgcctct 2220

aaattatctt ggtttggttt cagtgagaat ggtgaactta gttcctttga ttccaaggga 2280

atactgaggg tcttttctgg tcaattcggt ggaagctgga ttccaatatt tagttcaatc 2340

aaggcaagaa aatccgaaga tgaaagccac tgggtggtgg gcttagatgc taataatata 2400

ttctgcattc tatgcaagtc cccggagtcc tatccacagg tgatgcccaa acctgttttg 2460

acaatacttg agctgtcatt tcctcttgca tcatctgacc ttggtgccaa tagtttggaa 2520

actgaattca tgatgaggaa actgcatctc tcacagattc aaaagaaaat agaagaaatg 2580

gctgctttgg gtctggacac gattgcatta gatgatgaag cattcaacat ggaggctgca 2640

cttgaccggt gcatcttgag gctcatctcc agctgctgca atggtgataa gcttgtacga 2700

gctaccgagc ttgcaaaatt actaacactg gagaagtcaa tgaagggagc attgatgctt 2760

gttacacgct taaaacttcc catattgcaa gaaaggttca gtgccatact cgaggagatg 2820

atgctaaaca atgcaaaaat tgccaataca tctggtgttt tctccaatag taatacaaac 2880

tactcaccat caccagcgtt gagcactcaa gcagtcccac cagctaaggt tgtgcaaaat 2940

ggaaacagct tgaagttacc tacattgcct aaactgaatc ctgccgccca acgaagcaat 3000

ccaactgaat caaacaaggc agaggtagaa caagcagaca atttgaaaga aatcagtaca 3060

aaggtttcac ctgcacaaac tccgttagtt aaaattccaa aaaacagtga aatgggtgta 3120

aaaacgaaga aagataatga tggagcatca catgcaacta cagttgatca gaacccaaag 3180

ggaggcagtg gtcaggttgg ccttaaaaac aagagcgtcg atagctgcaa tggtgtacag 3240

cctcagcggc cagttaaccc cttcgcgaaa tcctcatcaa gcaaagaaca gccatcatcc 3300

ctttttgatt ccatcaagaa gatgaaggtc gaaaatgaga aggttgacaa agctaacagc 3360

aagaaggtga aagtgtaa 3378

<210>3

<211>1125

<212>PRT

<213> Artificial Sequence (Artificial Sequence)

<400>3

Met Ile Phe Gln Leu Arg Asn Ala Ile Val Lys His Ile Phe Leu Leu

1 5 10 15

Ser Leu Phe Leu Lys Ile Ser Leu Pro Tyr Leu Arg His Cys Gly Arg

20 25 30

Glu Ile Arg Cys Ile Phe Pro Ala Val Leu His Gln Lys Leu Pro Ala

35 40 45

Lys Arg Pro Phe Pro Arg Gly Gly Thr Arg Gln Asn Pro Pro Lys Cys

50 55 60

Leu Gln Pro Lys Pro His Pro Ala Tyr Arg Gln Ser Leu Glu Thr Pro

65 70 75 80

Asn Lys Phe Leu Asn Thr Arg Arg His His Ser Phe Ser Leu Val Gly

85 90 95

Glu Glu Ser Leu Val Val Val Val Gly Val Thr Met Lys Gly Arg Ala

100 105 110

Val Lys Leu Arg Glu Ala His Lys Ala Gly Ser Pro Val Phe Cys Ser

115 120 125

Val Ala Trp Gly Gln Gly Gly Gln His Val Val Thr Ala Ser Ala Ala

130 135 140

Asp Val Ala Ile Leu Ile His Asp Ala Ala Ala Val Ala Ala Ala Gly

145 150 155 160

Gly Arg Ser Ser Gly Ser Ala Ala Ala Ala Ala Leu Ser Thr Ile Arg

165 170 175

Leu His Lys Asp Gly Val Thr Ala Leu Ala Val Ala Pro Gly Ser Gly

180 185 190

Ala Ser Leu Ala Ser Gly Ser Ile Asp His Ser Val Lys Phe Cys Ser

195 200 205

Phe Pro Glu Gly Val Phe Gln Ser Asn Ile Ala Arg Phe Thr Leu Pro

210 215 220

Ile Arg Ser Leu Ala Phe Asn Lys Lys Gly Thr Leu Leu Ala Ala Ala

225 230 235 240

Gly Asp Asp Asp Gly Ile Lys Leu Ile Ala Thr Ile Asp Asn Thr Ile

245 250 255

Ser Lys Val Leu Lys Gly His Lys Gly Ser Val Thr Gly Leu Ser Phe

260 265 270

Asp Pro Arg Asn Asp Tyr Leu Ala Ser Ile Asp Thr Phe Gly Thr Val

275 280 285

Ile Phe Trp Asp Leu Cys Thr Gly Thr Glu Ala Arg Ser Leu Lys Arg

290 295 300

Ile Ala Pro Thr Phe Gly Ser Asp His Ser Ile Asn Asn Ala Leu Cys

305 310 315 320

Trp Ser Pro Asp Gly Gln Phe Leu Ala Val Pro Gly Leu Arg Asn Asn

325 330 335

Val Val Met Tyr Asp Arg Asp Thr Gly Glu Glu Val Phe Thr Leu Lys

340 345 350

Gly Glu His Glu Gln Pro Val Cys Ser Leu Cys Trp Ser Pro Asn Gly

355 360 365

Arg Tyr Leu Val Thr Ala Gly Leu Asp Lys Gln Val Leu Ile Trp Asp

370 375 380

Val Lys Ser Lys Gln Asp Val Glu Arg Gln Lys Phe Asp Glu Arg Ile

385 390 395 400

Cys Ser Leu Ala Trp Lys Pro Glu Ser Asn Ala Val Ala Val Ile Asp

405 410 415

Val Thr Gly Arg Phe Gly Ile Trp Glu Ser Val Ile Pro Ser Thr Leu

420 425 430

Lys Ser Pro Thr Glu Gly Ala Pro Asp Leu Asn Ser Thr Lys Val Pro

435 440 445

Leu Phe Asp Asp Glu Asp Asp Glu Glu Arg Pro Ser Thr Ser Gly Gly

450 455 460

Leu Asp Asp Asp Asp Asp Asp Glu Ser Leu Gly Glu Leu Gly Pro Phe

465 470 475 480

Asn His Lys Arg Leu Arg Arg Lys Ser Thr Tyr His Asp His Ser Asn

485 490 495

Gly Asp Ser Glu Asp Glu Asp Leu Ile Leu Gln Met Glu Ser Arg Lys

500 505 510

Arg Met Lys Asp Thr His Arg Asp Asn Lys Glu Val Ala Asp Lys Ala

515 520 525

Ile Gly Asp Ser Ala Thr Ser Val Arg Leu Val Thr Ala Arg Met Gln

530 535 540

Thr Ala Phe Gln Pro Gly Ser Thr Pro Pro Gln Pro Gly Lys Arg Asn

545 550 555 560

Phe Leu Ala Tyr Asn Met Leu Gly Ser Ile Thr Thr Ile Glu Asn Glu

565 570 575

Gly His Ser His Val Glu Val Asp Phe His Asp Thr Gly Arg Gly Pro

580 585 590

Arg Val Pro Ser Met Thr Asp Tyr Phe Gly Phe Thr Met Ala Ala Leu

595 600 605

Asn Glu Ser Gly Ser Val Phe Ala Asn Pro Cys Lys Gly Asp Lys Asn

610 615 620

Met Ser Thr Leu Met Tyr Arg Pro Phe Ser Ser Trp Ala Gly Asn Ser

625 630 635 640

Glu Trp Ser Met Arg Phe Glu Gly Glu Glu Val Lys Ala Val Ala Val

645 650 655

Gly Val Gly Trp Val Ala Ala Val Thr Thr Leu Asn Phe Leu Arg Ile

660 665 670

Phe Thr Glu Gly Gly Leu Gln Met His Ile Leu Ser Val Gly Gly Pro

675 680 685

Val Val Thr Ala Ala Gly His Gly Asp Gln Leu Ala Ile Val Ser His

690 695 700

Ala Ser Asp Cys Leu Ser Ser Gly Asp Gln Val Leu Asp Val Lys Val

705 710 715 720

Leu Lys Ile Ser Glu Cys Ala Gln Ser Leu Ser Ser Arg Leu Val Leu

725 730 735

Thr Pro Ala Ser Lys Leu Ser Trp Phe Gly Phe Ser Glu Asn Gly Glu

740 745 750

Leu Ser Ser Phe Asp Ser Lys Gly Ile Leu Arg Val Phe Ser Gly Gln

755 760 765

Phe Gly Gly Ser Trp Ile Pro Ile Phe Ser Ser Ile Lys Ala Arg Lys

770 775 780

Ser Glu Asp Glu Ser His Trp Val Val Gly Leu Asp Ala Asn Asn Ile

785 790 795 800

Phe Cys Ile Leu Cys Lys Ser Pro Glu Ser Tyr Pro Gln Val Met Pro

805 810 815

Lys Pro Val Leu Thr Ile Leu Glu Leu Ser Phe Pro Leu Ala Ser Ser

820 825 830

Asp Leu Gly Ala Asn Ser Leu Glu Thr Glu Phe Met Met Arg Lys Leu

835 840 845

His Leu Ser Gln Ile Gln Lys Lys Ile Glu Glu Met Ala Ala Leu Gly

850 855 860

Leu Asp Thr Ile Ala Leu Asp Asp Glu Ala Phe Asn Met Glu Ala Ala

865 870 875 880

Leu Asp Arg Cys Ile Leu Arg Leu Ile Ser Ser Cys Cys Asn Gly Asp

885 890 895

Lys Leu Val Arg Ala Thr Glu Leu Ala Lys Leu Leu Thr Leu Glu Lys

900 905 910

Ser Met Lys Gly Ala Leu Met Leu Val Thr Arg Leu Lys Leu Pro Ile

915 920 925

Leu Gln Glu Arg Phe Ser Ala Ile Leu Glu Glu Met Met Leu Asn Asn

930 935 940

Ala Lys Ile Ala Asn Thr Ser Gly Val Phe Ser Asn Ser Asn Thr Asn

945 950 955 960

Tyr Ser Pro Ser Pro Ala Leu Ser Thr Gln Ala Val Pro Pro Ala Lys

965 970 975

Val Val Gln Asn Gly Asn Ser Leu Lys Leu Pro Thr Leu Pro Lys Leu

980 985 990

Asn Pro Ala Ala Gln Arg Ser Asn Pro Thr Glu Ser Asn Lys Ala Glu

9951000 1005

Val Glu Gln Ala Asp Asn Leu Lys Glu Ile Ser Thr Lys Val Ser Pro

1010 1015 1020

Ala Gln Thr Pro Leu Val Lys Ile Pro Lys Asn Ser Glu Met Gly Val

1025 1030 1035 1040

Lys Thr Lys Lys Asp Asn Asp Gly Ala Ser His Ala Thr Thr Val Asp

1045 1050 1055

Gln Asn Pro Lys Gly Gly Ser Gly Gln Val Gly Leu Lys Asn Lys Ser

1060 1065 1070

Val Asp Ser Cys Asn Gly Val Gln Pro Gln Arg Pro Val Asn Pro Phe

1075 1080 1085

Ala Lys Ser Ser Ser Ser Lys Glu Gln Pro Ser Ser Leu Phe Asp Ser

1090 1095 1100

Ile Lys Lys Met Lys Val Glu Asn Glu Lys Val Asp Lys Ala Asn Ser

1105 1110 1115 1120

Lys Lys Val Lys Val

1125

<210>4

<211>21

<212>DNA

<213> Artificial Sequence (Artificial Sequence)

<400>4

gctacctagt caaattaatc g 21

<210>5

<211>21

<212>DNA

<213> Artificial Sequence (Artificial Sequence)

<400>5

gattaggcca agtaagtcca c 21

<210>6

<211>20

<212>DNA

<213> Artificial Sequence (Artificial Sequence)

<400>6

tgcatggtca cgttcctcat 20

<210>7

<211>20

<212>DNA

<213> Artificial Sequence (Artificial Sequence)

<400>7

attgcggagt gatgagagat 20

<210>8

<211>21

<212>DNA

<213> Artificial Sequence (Artificial Sequence)

<400>8

aaccaagcaa aagtcattgg a 21

<210>9

<211>21

<212>DNA

<213> Artificial Sequence (Artificial Sequence)

<400>9

cgagtaatat tttgggcgtc a 21

<210>10

<211>22

<212>DNA

<213> Artificial Sequence (Artificial Sequence)

<400>10

gcatcattag tcctggttag cg 22

<210>11

<211>20

<212>DNA

<213> Artificial Sequence (Artificial Sequence)

<400>11

gtggaactct ccactgctcc 20

<210>12

<211>20

<212>DNA

<213> Artificial Sequence (Artificial Sequence)

<400>12

gtgtagctat gggtaccatc 20

<210>13

<211>20

<212>DNA

<213> Artificial Sequence (Artificial Sequence)

<400>13

ggtctctcac tgtttctggt 20

<210>14

<211>20

<212>DNA

<213> Artificial Sequence (Artificial Sequence)

<400>14

gcgatcgggc aagttcaaaa 20

<210>15

<211>20

<212>DNA

<213> Artificial Sequence (Artificial Sequence)

<400>15

gggatctctg gaaaaaggac 20

<210>16

<211>20

<212>DNA

<213> Artificial Sequence (Artificial Sequence)

<400>16

tgtctgggaa gcttccaaca 20

<210>17

<211>20

<212>DNA

<213> Artificial Sequence (Artificial Sequence)

<400>17

ccaccaaagc cgactctata 20

<210>18

<211>20

<212>DNA

<213> Artificial Sequence (Artificial Sequence)

<400>18

gggtgattgg agatatgaca 20

<210>19

<211>20

<212>DNA

<213> Artificial Sequence (Artificial Sequence)

<400>19

agcatagcaa taatggccac 20

<210>20

<211>20

<212>DNA

<213> Artificial Sequence (Artificial Sequence)

<400>20

tttttgggga ataccctccc 20

<210>21

<211>20

<212>DNA

<213> Artificial Sequence (Artificial Sequence)

<400>21

ggtttggcac catgtggaaa 20

<210>22

<211>20

<212>DNA

<213> Artificial Sequence (Artificial Sequence)

<400>22

cataccttgc agtcctagaa 20

<210>23

<211>20

<212>DNA

<213> Artificial Sequence (Artificial Sequence)

<400>23

cgtggctagt ccatcaattc 20

<210>24

<211>42

<212>DNA

<213> Artificial Sequence (Artificial Sequence)

<400>24

cgaacgatag ccatggccat gatatttcag ctaagaaatg cg 42

<210>25

<211>40

<212>DNA

<213> Artificial Sequence (Artificial Sequence)

<400>25

ggtaggatcc actagtcact ttcaccttct tgctgttagc 40

<210>26

<211>37

<212>DNA

<213> Artificial Sequence (Artificial Sequence)

<400>26

ccatgattac gaattctgct atgccgttag gtagcac 37

<210>27

<211>44

<212>DNA

<213> Artificial Sequence (Artificial Sequence)

<400>27

cccttgctca ccatggtacc ctgaaatatc attgctaacc atca 44

<210>28

<211>44

<212>DNA

<213> Artificial Sequence (Artificial Sequence)

<400>28

caatgatatt tcagggtacc atgatatttc agctaagaaa tgcg 44

<210>29

<211>44

<212>DNA

<213> Artificial Sequence (Artificial Sequence)

<400>29

cccttgctca ccatggtacc cactttcacc ttcttgctgt tagc 44

<210>30

<211>37

<212>DNA

<213> Artificial Sequence (Artificial Sequence)

<400>30

cggtacccgg ggatcctgct atgccgttag gtagcac 37

<210>31

<211>38

<212>DNA

<213> Artificial Sequence (Artificial Sequence)

<400>31

ctcagatcta ccatgggaaa tatcattgct aaccatca 38

Claims (7)

1. The application of the rice MIT2 protein or the coding gene thereof or the biological material containing the coding gene thereof in any one or more of the following applications:

(1) the application in preparing transgenic plants;

(2) the application in rice germplasm resource improvement;

(3) the application in maintaining rice fertility;

(4) the application of the rice grain enlargement agent in enlarging rice grains;

(5) is an application for increasing the height of rice plants;

(6) the application in inhibiting rice tillering;

the amino acid sequence of the rice MIT2 protein is shown as SEQ ID number 3.

2. A rice MIT2 gene mutant gene is characterized in that a base T at the 2413 th position of a CDS (nucleotide sequence shown in SEQ ID No. 2) on the 8 th exon of a rice MIT2 gene is deleted.

3. An allelic variant of the rice MIT2 mutant gene according to claim 2, wherein the mutation site of the allelic variant occurs at a splicing site at the 3 'end of the first intron, the original AG is changed into AA, and the first two bases AG at the 5' end of the second exon are cut off.

4. A biomaterial containing the rice MIT2 gene mutant gene of claim 2 or the allelic mutant of claim 3, wherein the biomaterial is a plasmid, a vector, a host bacterium, or a transformed plant cell.

5. The use of any one or more of the rice MIT2 gene mutant gene of claim 2, the allelic mutant of claim 3, or the biomaterial of claim 4,

(1) the application in crop improvement breeding and seed production;

(2) the application in reducing the height of crop plants;

(3) the application in increasing the tillering number of rice;

(4) the application in reducing rice fertility;

(5) the application in cultivating rice with reduced grains;

(6) application in improving rice yield.

6. The use according to claim 5, wherein the reduction in plant height of the crop is to 50% -60% plant height compared to wild type.

7. The use according to claim 5, wherein the increase in rice tillering number is a 2-3 fold increase in tillering number compared to wild type.

Images